The relationship between elastase and lactoferrin in healthy, gingivitis and periodontitis sites

OBJECTIVES: To compare the relative amounts of elastase (primary polymorphonuclear leucocyte granule constituent) and lactoferrin (secondary PMN granule constituent) in the gingival crevicular fluid (GCF) of healthy, gingivitis and periodontitis sites.
DESIGN: This cross-sectional study looked at the two GCF constituents in three categories of disease status within the same subject.
MATERIALS AND METHODS: Patients with chronic adult periodontitis were screened and those exhibiting all three types of sites ie periodontally healthy, gingivitis and periodontitis sites were recruited (n = 10) and had GCF collected from the three sites. Lactoferrin and elastase were measured in eluates of GCF by enzyme-linked immunosorbent assay.
RESULTS: The absolute amount of lactoferrin measured in ng per 30 s samples was significantly lower in healthy and gingivitis sites as compared to periodontitis sites however this difference failed to reach significance when the concentration of lactoferrin in GCF was used as the analytical unit. No significant differences were found for elastase levels at any sites when expressed as either absolute amounts or concentrations. Secondary granule release, as evidenced by lactoferrin levels, occurs during cell migration and the process is independent of primary granule release, which is thought to correlate with PMN activation. The relationship between granule constituents in the samples showed significant differences, the highest lactoferrinlelastase ratio being at periodontitis sites (P < 0.001).
CONCLUSIONS These findings imply a change in the relative amounts of elastase and lactoferrin released at different disease level sites, with an almost 10-fold increase in the proportion of lactoferrin to elastase in periodontitis sites over healthy and gingivitis sites. This variation in the release by PMNs of primary and secondary granule constituents may indicate alterations in PMN function in different disease environments.     

Gingival crevicular fluid collagenase-2 (MMP-8) test stick for chair-side monitoring of periodontitis

BACKGROUND: A rapid chair-side test based on the immunological detection of elevated levels of collagenase-2 (matrix metalloproteinase-8, MMP-8) in gingival crevicular fluid (GCF) was developed to identify and monitor the course and treatment of adult periodontitis. METHODS: MMP-8 was determined in GCF from periodontitis (11 patients, 90 sites), gingivitis (10 patients, 58 sites) and healthy control (8 patients, 59 sites) sites (i) by a test stick incorporating monoclonal antibodies to two epitopes on MMP-8 and (ii) by measuring MMP-8 concentration by a quantitative immunofluorometric assay. Patients with adult periodontitis were treated by scaling and root planing (SRP) and received oral hygiene instructions. GCF MMP-8 testing and clinical measurements were done before and after SRP. RESULTS: MMP-8 GCF levels and chair-side test differentiated periodontitis from gingivitis and healthy control sites. MMP-8 GCF levels > 1 mg/l and positive chair-side test identified especially severe periodontitis sites. A positive and negative test stick result, the outcome of which was rapidly detectable in 5 mins, in GCF correlated well with MMP-8 immunofluorometric assay analysis from the collected GCF samples and the severity of periodontitis. Scaling and root planing reduced the MMP-8 levels in severe periodontitis sites with positive MMP-8 test and gingival probing pocket depth (PD) > 5 mm before treatment. The test stick result and the quantitative assay were discrepant in only 18 of the 207 sites tested, thus agreement was very good (kappa = 0.81). With a threshold of 1 mg/l MMP-8 activity the chair-side test provided a sensitivity of 0.83 and specificity of 0.96 (n = 207). CONCLUSION: The MMP-8 test can be used to differentiate periodontitis from gingivitis and healthy sites as well as to monitor treatment of periodontitis. A reduction in GCF MMP-8 levels and a change in test stick result provide a means to optimize patient control during maintenance of periodontal treatment.     

Plasminogen activator system in smokers and non-smokers with and without periodontal disease

BACKGROUND: The present study assessed levels of plasminogen activator (PA) system proteins in gingival crevicular fluid (GCF) and serum of chronic gingivitis, chronic periodontitis patients and periodontally healthy subjects and evaluated how smoking influenced these levels. METHODS: Twenty chronic gingivitis; 20 chronic periodontitis patients and 20 periodontally healthy volunteers were consecutively recruited according to the inclusion criteria so that exactly half of the subjects in each category were smokers. GCF samples from four sites together with serum samples were obtained from each subject. GCF levels of tissue type PA (t-PA), urokinase type PA (u-PA), PA inhibitor-1 (PAI-1) and PA inhibitor-2 (PAI-2) and serum concentrations of cotinine, u-PA and PAI-1 were analysed by enzyme-linked immunosorbent assay. RESULTS: The only statistically significant difference between smokers and non-smokers was a lower GCF PAI-2 concentrations in healthy smokers compared with healthy non-smokers (p<0.01). Gingivitis and periodontitis patients had higher GCF concentrations of PAI-2 than healthy subjects (p<0.002 and p<0.02 respectively). The ratio of u-PA:PAI-1 and t-PA:PAI-1 were significantly higher in GCF of smokers with periodontitis compared with "healthy" smokers, whereas the ratio of t-PA:PAI-2 was significantly lower in smokers with periodontal disease (p<0.05). CONCLUSIONS: GCF levels of the PA system proteins are increased in chronic gingivitis and periodontitis compared with healthy gingiva. Smoking had only subtle effects on the GCF PA system proteins with the exception of PAI-2, and the balance of activators and inhibitors. These findings suggest one mechanism whereby smoking may exert detrimental effects on the periodontal tissues.     

Smoking, periodontal disease and the role of the dental profession

Abstract:  Epidemiological investigations support a firm relationship between smoking and periodontal disease. The likely benefits of smoking cessation programmes are considerable for periodontal disease, cancers and nearly all chronic systemic diseases. The mechanisms by which smoking may influence the development and progression of periodontal disease are as yet unclear, but may include changes in the vasculature, the immune and inflammatory systems, tissue oxygenation and the healing processes. Unfortunately, although dental professionals have more opportunities to encourage smokers to quit (most people visit their dentist more frequently than their doctor), dentists claim that they are not well informed on this subject. The purpose of this review is to describe the evidence for a link between smoking and periodontal disease, the possible pathology induced by smoking on the periodontal tissues and its impact on therapy, and to outline the smoking cessation techniques that are currently available.     

Humoral immune response in early-onset periodontitis: influence of smoking

Sixty-five patients with generalised early-onset periodontitis (G-EOP) (age range 16-42 years, 32 smokers and 33 non-smokers) were assessed for antibody titres and avidity to a panel of five suspected periodontal pathogens (Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans, Prevotella intermedia, Treponema denticola and Bacteroides forsythus). Thirty-four of these patients were untreated (17 smokers and 17 non-smokers), and thirty-one were in the maintenance phase of periodontal therapy (15 smokers and 16 non-smokers). Previous studies have investigated the effect of smoking on IgG levels in periodontitis patients in the context of the more extensive periodontal destruction seen in smokers. Based on this literature our hypothesis was that smokers would have depressed serum IgG levels directed against recognised periodontal pathogens compared with non-smokers. Antibody titres were measured by ELISA deploying fixed whole cells as coating. The IgG response was detected with biotin-anti-human IgG and avidin-peroxidase; avidity was determined by elution with ammonium thiocyanate. Median titres to A. actinomycetemcomitans, P. intermedia and T. denticola were significantly lower in maintenance patient smokers (p= 0.02, 0.02 and 0.002 respectively) but not in untreated patients. Avidity to P. gingivalis was also lower in smoking maintenance patients (p = 0.003) but not in untreated patients. These findings may imply some interruption of immune maturation in smokers following periodontal treatment.     

Quadrant root planing versus same-day full-mouth root planing

OBJECTIVES: The aim of this study was to determine whether same-day full-mouth scaling and root planing (FM-SRP) and quadrant scaling and root planing (Q-SRP) resulted in variations in the systemic humoral immune response dynamics (antibody titres and avidity) during active treatment and 3 and 6 months post-therapy. MATERIALS AND METHODS: Forty patients with chronic periodontitis were recruited into this study. Subjects were randomised into two groups and received either scaling and root planing quadrant by quadrant at 2-weekly intervals (Q-SRP group) or same-day full-mouth scaling and root planing (FM-SRP group). Clinical measurements and serum samples were obtained at baseline and approximately 6 weeks after the last clinical intervention (R1) and 6 months after the initiation of therapy (R2). Furthermore, serum samples were obtained from each patient undergoing therapy (Q-SRP and FM-SRP) at 3 bi-weekly instances so as to determine the short-term effects of each session of scaling and root planing on the dynamics of the humoral immune response. Serum antibody titre was assayed by enzyme-linked immunosorbent assay (ELISA) and antibody avidity was measured by thiocyanate dissociation against five putative periodontal pathogens: Porphyromonas gingivalis; Actinobacillus actinomycetemcomitans; Prevotella intermedia; Treponema denticola and Bacteroides forsythus. RESULTS: Both therapies resulted in similar antibody titre reductions against the majority of the organisms tested and although there was a distinct trend for antibody avidity to increase following therapy, this was not found to be statistically significant, reflecting marked inter-individual variation. In addition, no evidence emerged from this study to support increased antibody titres following the active phases of both treatment approaches due to an inoculation effect. Nevertheless, significant short-term increases in antibody avidity to most test bacteria were noted for both treatment strategies. CONCLUSION: Both therapies were associated with a reduction in antibody titres and an increase in the binding ability or avidity of antibodies, but there was a marked inter-subject variability and statistical significance was reached for only some of the test bacteria. No significant differences in the humoral antibody dynamics were found between the two treatment approaches.     

The systemic immune response is more prominent than the mucosal immune response in the pathogenesis of periodontal disease

BACKGROUND, AIM: The diseased periodontium appears to express features of a systemic and a mucosal immune response. Our aims were to determine differences in immunoglobulin expression between gingivitis and periodontitis lesions and to ascertain whether immune and inflammatory cells were recruited into the diseased periodontium by the mucosal addressin adhesion molecule (MAdCAM-1). METHODS: In situ hybridization and immunohistochemistry were used to detect the expression of chemokines, adhesion molecules and immunoglobulins in tissue sections of gingival and granulation tissues excised from periodontitis-affected sites and of healthy tissue and gingivitis-affected tissue excised during crown-lengthening procedures. RESULTS: Greater numbers of plasma cells were observed in periodontitis gingival/granulation tissue lesions compared with gingivitis lesions. While IgA1 were predominant in all lesions, IgA2 and J-chain expressing plasma cells were present in increased proportions in gingival tissues compared with granulation tissue. Intracellular adhesion molecule-1 (ICAM-1) was higher in periodontitis than in gingivitis and interleukin-8 mRNA was higher in lesions with a pronounced neutrophil infiltrate. Vascular cell adhesion molecule-1 (VCAM-1) localized to the deep connective tissue and indicated the presence of a systemic type of immune response in this region. Periodontal tissues (n=71 biopsies) did not appear to express MAdCAM-1, in positive control sections of small intestine where it was detected. CONCLUSION: Overall, the systemic-type immune response is predominant, and although the mucosal immune response is minor and limited to the superficial tissues it may have an important role in the host defense to periodontal pathogens.     

Changes in subgingival microflora and humoral immune response following periodontal therapy

OBJECTIVES: To investigate the effect of scaling and root planing (SRP) on the microflora and humoral immune response in adult periodontitis. MATERIALS & METHODS: Clinical measurements, subgingival plaque samples, gingival crevicular fluid and sera were taken from 4 sites in 28 adult periodontitis patients before and after SRP. Polymerase chain reaction was used to determine the presence of A. actinomycetemcomitans, P. gingivalis, B. forsythus, P. intermedia, and T. denticola. ELISA was used to investigate the systemic and local antibody titres to these organisms, and thiocyanate dissociation for the determination of serum antibody avidity. RESULTS: SRP produced a good clinical improvement. On a subject basis there was little significant change in the microflora. However, on a site basis, there were significant reductions in P. intermedia, B. forsythus and T. denticola. There was little change in systemic and local antibody titres following SRP, although there was a significant reduction in antibody avidity to P. gingivalis and P. intermedia CONCLUSION: Post-therapy clinical improvement was associated with a reduction in bacterial prevalence, but statistical significance was only reached at a site level and this microbial reduction was not significant for all organisms. No significant post-therapy effects on the humoral immune response were noted other than a reduced antibody avidity to P. gingivalis and P. intermedia. The lack of a clear pattern in the humoral immune response may reflect a failure of the host response to produce adequate levels of biologically functional antibodies, and complex interactions between the subgingival flora and the host response.     

Crevicular fluid and serum IgG subclasses and corresponding mRNA expressing plasma cells in periodontitis lesions

Recent reports suggest that specific serum IgG subclasses are a feature of several forms of periodontitis. GCF antibodies are both serum-derived and locally produced by the abundant plasma cells of the diseased periodontal tissue. Previous work has shown that crevicular fluid (GCF) levels of IgG may be reduced in active and deep periodontal pockets when compared to other sites in chronic periodontitis patients (7). These findings, and more recent findings for IgA levels in GCF (5), suggest that GCF immunoglobulins may indicate "high risk" sites for periodontitis. In these studies, the relative distribution of IgG isotypes was not investigated, nor was the relative contribution of local and serum antibodies to the GCF immunoglobulin profile. Therefore, more precise investigation of the tissue distribution of local gingival IgG subclass producing plasma cells and their protein levels in the GCF from the same sites and in serum, was undertaken.